[Histonet] Triton and Secondary Mouse

From:"Pixley, Sarah \(pixleysk\)"

 Dear Sohail on Triton:
For whole mounts we have used 1% Triton (in PBS or PB with sodium azide)
in primary, secondary and ABC (no sodium azide in ABC!) reagents. We
incubated with primary antibodies for up to one week, and secondary and
ABC each for overnight, with longer and more washes than normal between
each step. We use 0.2% for all other IHC. (We are not a histotechnology
lab, though, so I don't know if this meets those standards).  

Dear Gudrun on mouse IHC:
When we use a mouse primary antibody on mouse tissues, it is luck if the
primary will work properly or not. For a secondary, we use a
biotinylated secondary mouse antibody from Jackson that is absorbed
against rat tissues. This gives us a very clean background, cleaner than
the regular anti-mouse antibody. Theoretically, the secondary should
only be attaching to cells of the blood system, like macrophages and
tissue lymphocytes. But there is other background. My suggestion is to
try the "rat-absorbed" version of anti-mouse.

Best wishes
Sarah Pixley

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